Biophysical Chemistry, Center for Molecular Protein Science, Department of Chemistry , Lund University , P.O. Box 124, SE-22100 Lund , Sweden.
Biochemistry and Structural Biology, Center for Molecular Protein Science, Department of Chemistry , Lund University , P.O. Box 124, SE-22100 Lund , Sweden.
J Am Chem Soc. 2019 Nov 27;141(47):18649-18652. doi: 10.1021/jacs.9b07952. Epub 2019 Nov 12.
Amyloid fibril formation is a hallmark of neurodegenerative disease caused by protein aggregation. Oligomeric protein states that arise during the process of fibril formation often coexist with mature fibrils and are known to cause cell death in disease model systems. Progress in this field depends critically on development of analytical methods that can provide information about the mechanisms and species involved in oligomerization and fibril formation. Here, we demonstrate how the powerful combination of diffusion NMR and multilinear data analysis can efficiently disentangle the number of involved species, their kinetic rates of formation or disappearance, spectral contributions, and diffusion coefficients, even without prior knowledge of the time evolution of the process or chemical shift assignments of the various species. Using this method we identify oligomeric species that form transiently during aggregation of human superoxide dismutase 1 (SOD1), which is known to form misfolded aggregates in patients with amyotrophic lateral sclerosis. Specifically, over a time course of 42 days, during which SOD1 fibrils form, we detect the disappearance of the native monomeric species, formation of a partially unfolded intermediate in the dimer to tetramer size range, subsequent formation of a distinct similarly sized species that dominates the final spectrum detected by solution NMR, and concomitant appearance of small peptide fragments.
淀粉样纤维的形成是由蛋白质聚集引起的神经退行性疾病的一个标志。在纤维形成过程中出现的低聚物蛋白状态常常与成熟纤维共存，并已知在疾病模型系统中导致细胞死亡。该领域的进展取决于分析方法的发展，这些方法可以提供关于寡聚化和纤维形成过程中涉及的机制和物种的信息。在这里，我们展示了扩散 NMR 和多线性数据分析的强大组合如何能够有效地分离出所涉及的物种数量、它们的形成或消失的动力学速率、光谱贡献和扩散系数，即使没有过程的时间演变或各种物种的化学位移分配的先验知识。使用这种方法，我们鉴定了人超氧化物歧化酶 1（SOD1）聚集过程中形成的瞬态低聚物物种，已知 SOD1 在肌萎缩侧索硬化症患者中形成错误折叠的聚集体。具体来说，在 42 天的时间过程中，SOD1 纤维形成，我们检测到天然单体物种的消失，二聚体到四聚体大小范围内部分展开的中间体的形成，随后形成一个独特的类似大小的物种，主导最终通过溶液 NMR 检测到的光谱，同时出现小的肽片段。
